Desmogleins are members of the cadherin superfamily of cell adhesion molecules. Along with desmocollins (Dscs), desmogleins (Dsgs) make up the adhesive core of the desmosome through their extracellular domains, and anchor intermediate filaments to the plasma membrane through interactions between their cytoplasmic domains and armadillo/plakin proteins. Autoantibodies directed against desmogleins are causative in the blistering diseases known as pemphigus; however, the molecular mechanism underlying pathogenesis in pemphigus is poorly understood. Furthermore, there are three Dsg genes and three Dsc genes, expressed in a differentiation-dependent manner in the epidermis, and it is unclear why multiple genes are required. To understand the underlying basis of human autoimmune and inherited disorders targeting the desmogleins, it will be crucial to determine the functions of desmosomal cadherins as well as how the adhesive complex is assembled, maintained and regulated. Our aims for the next funding period are: 1) To determine the molecular basis of desmosomal cadherin-mediated adhesion and pairing preferences for Dscs and Dsgs by using a tetracycline-regulatable system in which adhesive function can be reconstituted in L929 cells, and to develop this system into a test for the pathogenic activity of pemphigus antibodies, 2) To determine how Dsgs/Dscs collaborate with armadillo proteins to form distinct, differentiation-specific protein complexes using a combination of in vitro and high resolution structural studies to establish the identity and affinity of desmosomal cadherin-armadillo binding partners, and cellular reconstitution techniques to determine how interactions translate into structurally distinct desmosomes, 3) To determine how desmosomal cadherins and their associated proteins are coordinated spatially and temporally during their assembly into keratinocytes by time lapse analysis of living cells using fluorescently-labeled probes, and 4) To determine the function of Dsg1 and biological significance of Dsg1 as a caspase substrate for keratinocyte adhesion, differentiation and apoptosis by defining UV-dependent Dsg1 processing events that occur in vitro and in cultured keratinocytes and by defining the roles of wild type Dsg1 its cleavage products and cleavage resistant forms during differentiation in cultured human keratinocytes and transgenic mice.